Neisseria gonorrhoeae remains a critically important STD pathogen, and development of a vaccine to prevent this infection is a national priority. The long-term goals of our work are to contribute to this objective. We have developed a model of experimental gonococcal urethritis in male volunteer subjects which closely resembles natural infection. To date we have studied more than 80 subjects. We plan to use this model to identify bacterial virulence factors as to better understand phenotypic variation of gonococci in vivo, and to test vaccines. In the current Application we describe experiments using isogenic mutants of gonococcal strain F1090 generated with no new antibiotic markers. The following isogenic mutants will be generated to determine whether particular virulence facts are required to cause urethral infection: i) "pan-Opa" protein mutant, which may not attach or invade epithelial cells as well as control organisms; ii) a tbpA-tbpB mutant, which cannot generate outer membrane proteins gonococci require to use iron bound to transferrin; iii) a catalase mutant, which would not be expected to protect gonococci from H2O2 and/or oxidative neutrophil attach=k, and iv) lipooligosaccharide mutants which cannot undergo sialylation, or generate the repertoire of sugars observed in vivo. We have calculated the number of subjects required for each of these studies, so that difference between the isogenic mutant and the control can achieve statistical significance. We believe that that this work will lead to a better understanding of gonococcal pathogenesis, and help to prepare for the use of this model for vaccine development and testing.